Stereochemistry, the study of three dimensional structure of molecules, is essential to the understanding organic chemistry, biochemistry, and biological systems. For example, biological systems are exquisitely selective and they often discriminate between molecules with subtle stereochemical differences. There are many forms of stereochemical differences such as structural isomers or constitutional isomers, wherein the isomers consist of the same molecular formula, but differ in their bonding sequences, i.e. the atoms are connected differently. Another form of stereochemical difference is known as stereoisomers, wherein the isomers have the same bonding sequence, but they differ in the orientation of their atoms in space. One form of stereoisomers is known as enantiomers. Enantiomers are pairs of compounds that are non-superimposable mirror images, which may also be referred to as chiral compounds. An analogous example of a chiral compound is the left and the right hand. The left and the right hand are mirror images but are non-superimposable on one another. Chiral compounds are important to many biological functions. Enzymatic reactions, drug metabolism, pharmaceutically active sites, and many other biological functions are only reactive to one isomer or enantiomeric form and inert or toxic the other form. Thus, in the pharmaceutical industry alone there is a strong need for an effective means to separate the stereoisomers.
The separation and detection of enantiomeric mixture or trace level enantiomeric impurity from the major enantiomer into individual optical isomers is one of the most challenging problems in analytical chemistry. This has important considerations in many areas of science, in particular the pharmaceutical and agricultural industries. For example, according to according to US Food and Drug Administration (FDA) and International Conference on Harmonization (ICH) Guidance for Industry Q3A Impurities in New Drug Substances, a chiral assay with a LOD of 0.1% enantiomeric impurity is mandatory for later stage of drug development. Separation of optical isomers is often very challenging due to the need of considerable time, effort, and expense.
High performance liquid chromatography and capillary electrophoresis are common techniques currently used to separate chiral compounds. Another separation technique includes reversible complexes formed of metal ions and chiral complexing agents, also known as ligand-exchange-chromatography. This technique involves multicomponent complexes containing a central metal ion and two chelating chiral molecules. The enantiomers are separated by using chiral mobile phase additives or by using a chiral stationary phase. Additionally, researchers have used micelles in chiral compound separation.
Micelles are typically surfactants. Micelles usually comprise both hydrophilic and hydrophobic groups which associate with one another in polar solvents such as water to form dynamic aggregates. A micelle typically takes roughly the shape of a sphere, a spheroid, an ellipsoid, or a rod, with the hydrophilic groups on the exterior and the hydrophobic groups on the interior. The hydrophobic interior provides, in effect, a hydrophobic liquid phase with solvation properties differing from those of the surrounding solvent. As a result, micelles have been used in applications of chiral recognition and separation. Chiral surfactants have been used to form micelles having distinct chiral properties. The resulting chiral microenvironment has been shown to exhibit selective interactions with different enantiomers in solution. However, micelle separation often results in heat, which could promote degradation in the compounds. Another disadvantage with traditional micelle chiral compound separation is that they have slow mass transfer rates and often times form troublesome emulsion systems. Conventional micelles include alkenoxy based amino acid surfactant with carboxylate head group, which often causes precipitation in solution at various pHs. This is mainly because the utility of alkenoxy amino acid-based surfactants is somewhat limited by the carboxylate head groups whose pH chemistry limits the electrophoretic mobility and solubility of the molecular micelles at pHs below 5.5.
Not only is separation of isomers important, but analysis and characterization of the isomers is equally important. The use of conventional chiral surfactant (above its critical micelle concentration) for capillary electrophoresis (CE) with mass spectrometry (MS) of optical isomers is not trivial. This is due to nonvolatility and high surface activity of the unpolymerized micelles. In addition, the use of low-molecular weight surfactant monomers provides a very unstable electrospray due to the large background signal generated from the dissociation of a micelle, which ultimately leads to the fouling of the ionization source and limits the sensitivity of the electrospray ionization-mass spectrometry (ESI-MS) applications. Finding a low molecular weight chiral selector that is MS-compatible is very difficult. Most applications concentrate on the use of partial-filling (PF)-CE-MS in which only part of the capillary is filled with micelle containing buffers. While this approach can be useful, it results in significant deterioration in chiral resolution.
Thus, there is a need for a more efficient and cost effective means for separation of optical isomers. In addition, because during the later stages of the drug development, sensitive analysis of the undesired optical isomer also is warranted. Thus, there is a continuing and growing need for improved separation and detection methods for chiral analysis.